Biology subjects

Sales, D.

Publications and source records attributed to Sales, D..

3 recordsLinked to original sources

bioRxiv2020

Transcriptomics, metabolomics and lipidomics of chronically injured alveolar epithelial cells reveals similar features of IPF lung epithelium

The current hypothesis suggests that Idiopathic pulmonary fibrosis (IPF) arises as a result of chronic injury to alveolar epithelial cells and aberrant activation of multiple signaling pathways. Dysfunctional IPF lung epithelium manifests many hallmarks of aging tissues, including cellular senescence, mitochondrial dysfunction, metabolic dysregulation, and loss of proteostasis. Unfortunately, this disease is often fatal within 3-5 years from diagnosis, and there is no effective treatment. One of the major limitations to the development of novel treatments in IPF is that current models of the disease fail to resemble several features seen in elderly IPF patients. In this study, we sought to develop an in vitro epithelial injury model using repeated low levels of bleomycin to mimic the phenotypic and functional characteristics of the IPF lung epithelium. Consistent with the hallmarks of the aging lung epithelium, we found that chronic-injured epithelial cells exhibited features of senescence cells, including an increase in {beta}-galactosidase staining, induction of p53 and p21, mitochondrial dysfunction, excessive ROS production, and proteostasis alteration. Next, combined RNA sequencing, untargeted metabolomics, and lipidomics were performed to investigate the dynamic transcriptional, metabolic, and lipidomic profiling of our in vitro model. We identified that a total of 8,484 genes with different expression variations between the exposed group and the control group. According to our GO enrichment analysis, the down-regulated genes are involved in multiple biosynthetic and metabolic processes. In contrast, the up-regulated genes in our treated cells are responsible for epithelial cell migration and regulation of epithelial proliferation. Furthermore, metabolomics and lipidomics data revealed that overrepresented pathways were amino acid, fatty acid, and glycosphingolipid metabolism. This result suggests that by using our in vitro model, we were able to mimic the transcriptomic and metabolic alterations of those seen in the lung epithelium of IPF patients. We believe this model will be ideally suited for use in uncovering novel insights into the gene expression and molecular pathways of the IPF lung epithelium and performing screening of pharmaceutical compounds.

cell biology↗

bioRxiv2020

Impaired HSF1 transactivation drives proteostasis collapse and senescent phenotype of IPF lung fibroblast.

Loss of proteostasis and cellular senescence are key hallmarks of aging. Recent studies suggest that lung fibroblasts from idiopathic pulmonary fibrosis (IPF) show features of cellular senescence, decline in heat shock proteins (HSPs) expression and impaired protein homeostasis (proteostasis). However, direct cause-effect relationships are still mostly unknown. In this study, we sought to investigate whether the heat shock factor 1 (HSF1), a major transcription factor that regulates the cellular HSPs network and cytoplasmic proteostasis, contributes to cellular senescence in lung fibroblasts. We found that IPF lung fibroblasts showed an upregulation in the expression of various cellular senescence markers, including {beta}-galactosidase activity (SA-{beta}-gal) staining, the DNA damage marker {gamma}H2Ax, the cell cycle inhibitor protein p21, and multiple senescence-associated secretory proteins (SASP), as well as upregulation of collagen 1a1, fibronectin and alpha-smooth muscle actin (-SMA) gene expression compared with age-matched controls. These changes were associated with impaired proteostasis, as judged by an increase in levels of p-HSF1ser307 and HSF1K298 sumo, downregulation of HSPs expression, and increased cellular protein aggregation. Similarly, lung fibroblasts isolated from a mouse model of bleomycin-induced lung fibrosis and mouse lung fibroblast chronically treated with H2O2 showed downregulation in HSPs and increased in cellular senescence and SASP markers. Moreover, sustained pharmacologic activation of HSF1 increased the expression of HSPs, reduced cellular senescence markers and effectively reduced the expression of pro-fibrotic genes in IPF fibroblast. Our data provide evidence that the HSF1-mediated proteostasis is important for driving lung fibroblasts toward cellular senescence and a myofibroblast phenotype. We postulate that enhancing HSF1 activity could be effective in the treatment of lung fibrosis.

cell biology↗

bioRxiv2018

Proteomic Profile of TGF-β1 treated Lung Fibroblasts identifies Novel Markers of Activated Fibroblasts in the Silica Exposed Rat Lung

We performed liquid chromatography-tandem mass spectrometry (LC-MS/MS) on control and TGF-{beta}1-exposed rat lung fibroblasts to identify proteins differentially expressed between cell populations. A total of 1648 proteins were found to be differentially expressed in response to TGF-{beta}1 treatment and 196 proteins were expressed at [≥] 1.2 fold relative to control. Guided by these results, we next determined whether similar changes in protein expression were detectable in the rat lung after chronic exposure to silica dust. Of the five proteins selected for further analysis, we found that levels of all proteins were markedly increased in the silica-exposed rat lung, including the proteins for the very low density lipoprotein receptor (VLDLR) and the transmembrane (type I) heparin sulfate proteoglycan called syndecan 2 (SDC2). Because VLDLR and SDC2 have not, to our knowledge, been previously linked to the pathobiology of silicosis, we next examined whether knockdown of either gene altered responses to TGF-{beta}1 in MRC-5 lung fibroblasts. Interestingly, we found knockdown of either VLDLR or SDC2 dramatically reduced collagen production to TGF-{beta}1, suggesting that both proteins might play a novel role in myofibroblast biology and pathogenesis of silica-induced pulmonary fibrosis. In summary, our findings suggest that performing LC-MS/MS on TGF-{beta}1 stimulated lung fibroblasts can uncover novel molecular targets of activated myofibroblasts in silica-exposed lung.\n\nHighlightsWe identified 196 proteins differentially expressed between control and TGF-{beta}1 treated fibroblastsby LC-MS/MS.\n\nSeveral proteins identified by LC-MS/MS were also found to be differentially expressed in whole lung tissues and isolated fibroblasts after chronic exposure to silica dust, including the very low density lipoprotein receptor (VLDLR) and the transmembrane type I heparan sulfate proteoglycan called syndecan 2\n\nKnockdown of SDC2 or VLDLR markedly inhibited collagen production in MRC-5 fibroblasts, suggesting a novel pathogenic role for these proteins in myofibroblast biology.

pharmacology and toxicology↗